摘要
目的从全人源单链噬菌体抗体库中筛选出抗前列腺特异性膜抗原(PSMA)特异性单链抗体并进行免疫活性鉴定。方法以合成的PSMA多肽为抗原,经过五轮吸附-洗脱-扩增,从单链噬菌体抗体库中筛选出特异性抗PSMA噬菌体抗体,ELISA检测其抗原结合能力,并对特异性较强的克隆提取质粒,表达可溶性抗体。WesternBlotting和免疫组织化学检测其抗原结合性,非竞争ELSIA法检测其亲和常数。结果从单链噬菌体抗体库中筛选出的噬菌体抗体,经ELISA鉴定为抗PSMA的特异性噬菌体抗体。抗PSMA可溶性抗体相对分子质量约为3.0×104,与PSMA特异性结合,其亲和常数约为5.077×106L/mol。结论所得全人源抗PSMA单链抗体保留完整抗体分子结合抗原的特异性,免疫原性弱,是肿瘤导向治疗的理想载体。
Objective To screen and identify a human single-chain variable fragment（scFv） antibody against prostate specific membrane antigen（PSMA） from a human scFv antibody library. Methods Using a synthetic PSMA peptide as the coating antigen, the antibody library was screened by five rounds of combining-eluting-amplification. The phage antibody against PSMA with high specificity was screened out from the human scFv antibody library and its binding abil- ity to the antigen was tested by ELISA. The soluble antibody was produced by plasmids extracted from highly specific clones, whose binding ability to PSMA was further identified by Western blot and immunohistochemistry. The affinity constant of the soluble antibody was measured by non-competitive ELISA. Results The screened phage antibody was specific for PSMA by ELISA. The soluble antibody was also specific for PSMA, its molecular weight was about 30 kD by SDS-PAGE and its affinity constant was about 5.077×10^6L/mol. Conclusions The screened scFv antibody is specific and has low immunogenicity. It can be further used in the target treatment of malignant tumors.
出处
《山东大学学报（医学版）》
CAS
北大核心
2011年第1期53-56,61,共5页
Journal of Shandong University:Health Sciences
基金
江苏省135医学重点人才基金资助项目(RC2002076)
关键词
前列腺特异性膜抗原
噬菌体抗体库
噬菌体抗体
亲和常数
Prostate specific membrane antigen
Phage antibody library
Phage antibody
Affinity constant
